The heart has a tremendous capacity for ATP generation which allows it to function as an efficient pump throughout the life of the organism. The adult myocardium uses either fatty acid (FA) and/or glucose oxidation as its main energy sources. Under normal conditions, the adult heart derives most of its energy through oxidation of fatty acids in mitochondria.
Cells of the myocardium have the ability to switch between carbohydrate glycolysis and the Krebs cycle and to fat fuel sources so that ATP production is maintained at a constant rate under diverse physiological and dietary conditions. This metabolic and fuel selection flexibility is important for normal cardiac function. Although cardiac energy conversion capacity and metabolic flux is modulated at many levels, one important mechanism of regulation occurs at the level of gene expression. The expression of genes involved in multiple energy transduction pathways is dynamically regulated in response to developmental, physiological, and pathophysiological cues.
The genes involved in these key energy metabolic pathways are transcriptionally regulated by members of the nuclear receptor superfamily, specifically the fatty acid-activated peroxisome proliferator-activated receptors (PPARs) and the nuclear receptor coactivator, PPARγ coactivator-1α (PGC-1α), as well as the estrogen receptor-related protein ERRα, ERRβ and ERRγ and their activators PGR-1 and PERC. The dynamic regulation of the cardiac PPAR/PGC-1 complex in accordance with physiological and pathophysiological states is described in more detail below.
PGC-1α is a PPARγ coactivator, linked to adaptive thermogenesis in brown adipose. Two structurally related proteins, PGC-1β and PARC, have been cloned and appear to be involved in regulating energy metabolic pathways. The tissue-specific and inducible nature of PGC-1α expression suggests its involvement in the dynamic regulation of cellular energy yielding metabolic processes, including mitochondrial biogenesis and oxidation, hepatic gluconeogenesis, and skeletal muscle glucose uptake. PGC-1α is selectively expressed in highly oxidative tissues such as heart, skeletal muscle, brown adipose, and liver. In the heart PGC-1α expression increases sharply at birth. This coincides with a perinatal shift from glucose metabolism to fat oxidation. PGC-1α activity and expression levels are also known to be induced by cold exposure, fasting, and exercise; stimuli known to promote oxidative metabolism. Forced expression of PGC-1 in cardiac myocytes in culture induces expression of nuclear and mitochondrial genes involved in multiple mitochondrial energy-transduction/energy-production pathways, increases cellular mitochondrial number, and stimulates coupled respiration. Signaling pathways associated with these stimuli, including p38 MAP kinase, β-adrenergic/cAMP, nitric oxide, AMP kinase, and Ca2-calmodulin kinase, activate PGC-1α and its downstream target genes either by increasing PGC-1α expression or its transactivation function.
These metabolic and structural changes can result in dilated cardiomyopathy and diastolic dysfunction in the heart. Interestingly, mitochondrial proliferation is reversible and the cardiomyopathy can be rescued upon reduction in transgene expression. This suggests that, in addition to serving as an activator of cellular fatty acid metabolism through PPARs, PGC-1α is linked to the mitochondrial biogenesis. Therefore, PGC-1α appears to serve as a master modulator of oxidative energy metabolism and responds to changes in the cellular energy status.
Evidence is emerging that the estrogen-related receptor (ERR) family of orphan Nuclear Receptors function as PGC-1-activated regulators of cardiac and skeletal muscle energy metabolism. There are three members of the ERR family: ERRα, ERRβ, and ERRγ. ERRα and ERRγ expression is elevated in adult tissues that rely primarily on mitochondrial oxidative metabolism for ATP production, such as heart and slow twitch skeletal muscle. ERRα expression dramatically increases in heart after birth, in parallel with the global upregulation of enzymes involved in cellular fatty acid uptake and mitochondrial oxidation. Recently, ERRα and ERRγ were identified as novel partners for the PGC-1 family of coactivators. This functional relationship between ERR isoforms and PGC-1α have stimulated interest in the role of ERRs in energy metabolism.
Deletion of the ERRα gene reveals a tissue-specific role for ERRα in constitutive regulation of lipid metabolism. White adipose mass is decreased in ERRα-/- mice coincident with decreased adipocyte size and lipid synthesis rates. In contrast, ERRα likely plays a role in lipid catabolism in heart, consistent with its functional interaction with PGC-1α. ERRα-/- mice, which do not display an overt cardiac phenotype, exhibit a compensatory increase in cardiac PGC-1α and ERRγ expression. These results suggest that ERR isoforms contribute to constitutive expression of fatty acid metabolic genes in heart. However, the metabolic effects of changes in gene expression remain unknown.
Gene expression profiling in cardiac myocytes that overexpress ERRα are being used to identify cardiac ERRα target genes. ERRα activates genes involved in energy production pathways, including cellular fatty acid uptake (LPL, CD36/FAT, H-FABP, FACS-1), β-oxidation (MCAD, VLCAD, LCHAD), and mitochondrial electron transport/oxidative phosphorylation (cytochrome c, COXIV, COXVIII, NADH ubiquinone dehydrogenase, flavoprotein-ubiquinone oxidoreductase, ATP synthase β). ERRα also increases palmitate oxidation rates in cardiac myocytes. Activation of β-oxidation enzymes genes by ERRα involves the PPARα signaling pathway. ERRα directly activates PPARα gene expression, and ERRα-mediated regulation of MCAD and M-CPT I is abolished in cells derived from PPARα-/- mice. ERRα is also now known to be involved in the PGC-1α regulation of mitochondrial biogenesis. It is known to mediate PGC-1α activation of the NRF pathway through regulation of the Gapba gene, which encodes a subunit of the NRF-2 complex and directly activates genes involved in mitochondrial oxidative metabolism at the level of transcription. ERRα with its coactivator PGC-1α activates the MCAD, cytochrome c, and ATP synthase β gene promoters. Collectively, these results identify ERRα as a regulator of cardiac oxidative energy metabolism through its involvement in the PGC-1 regulatory circuit. However, the precise biological roles of ERRs in heart have not been identified.
The nuclear receptor ERRγ (estrogen related receptor gamma) is highly expressed in heart, skeletal muscle, kidney, and brain, as well as in the developing nervous system. The expression of the coactivators PGC-1α and PGC-1β in mammalian cells potently augmented transcriptional activation by ERRγ. The constitutive activation function 2 (AF-2) of the orphan receptor is important for the synergistic enhancement. Functional receptor truncation analysis has been used to identify an additional amino-terminal activation function, specific for the ERRγ2 isoform and PGC-1α. In vitro experiments showed a direct interaction of ERRγ with both coactivators. These findings are consistent with the hypothesis that distinct regulatory functions for PGC-1α and PGC-1β as tissue-specific coactivators for ERRγ. Nevertheless, more studies are needed to further define these functions.
Cardiac-specific overexpression of PGC-1 in transgenic mice results in uncontrolled mitochondrial proliferation in cardiac myocytes leading to loss of sarcomeric structure and a dilated cardiomyopathy. Thus, PGC-1 is an important regulatory molecule in the control of cardiac mitochondrial number and function in response to energy demands.
Most, if not all of these regulatory pathways involve phosphorylation of intermediates in a signaling pathway. Inhibition of phosphorylation, such as by the action of various kinase inhibitors, affects these signaling pathways causing alterations in fatty acid metabolism which can cause organ toxicity, including cardiotoxicity. Many new anti-cancer drugs are kinase inhibitors and are accompanied by toxicity. Thus, methods are needed for identifying whether drugs may be accompanied by toxic effects and whether the toxic effects are likely to occur in a patient. Methods are also needed for avoiding toxic effects of these inhibitors while maintaining their potency against the phosphorylated receptor targets.